Xtt cell viability kit

SAHA cytotoxicity was assessed with a XTT Cell Viability Kit (30007, Biotium®) after a 24 h-treatment according to the manufacturer’s instructions. Briefly, 100 µL of activated XTT solution were added on each well. After 5 h of incubation, absorbance was measured at 450 nm and 630 nm.

Cell viability (metabolic activity) was detected using a commercially available colorimetric XTT assay, following manufacture’s protocol (XTT Cell Viability Kit, Biotium, CA, USA). Briefly, cells were cultured on 96-well plates, at a concentration of 15,000 cells in 100 µL, per well. The cells were treated or not with CAR for three days before irradiation, unirradiated cells were used as controls. Cells were irradiated and two hours after irradiation 25 μl XTT solution was added to control and irradiated cells, plates were incubated for 2 hours after XTT addition. The cell viability was determined by measuring the absorbance of the wells at a wavelength of 450 nm, background absorbance was measured at a wavelength of 630 nm. Background absorbance was subtracted from signal absorbance to obtain normalized values. Results were expressed as the percentage of XTT reduction, taking the absorbance of control cells as 100%.

Viability assays were performed with an XTT Cell Viability Kit (Biotium, Hayward, CA, USA) according to the manufacturer's protocol. 5000 cells were seeded in sixplicate, and treated with increased doses of GSK-J4 (Sigma-Aldrich, St Louis, MO, USA) for 24 h, 48 h or 72 h. After 2 h of XTT incubation, cell viability was determined by measuring the absorbance signal at 450 nm with a Multiskan™ GO Microplate Spectrophotometer (ThermoFisher Scientific). Viable cells were presented as a percentage of the untreated cell control, and IC₅₀ was determined by linear interpolation between concentrations just above and below 50% inhibition in the response dose curve. IC₅₀ was calculated using the formula: EXP(LN(conc > 50%)-((signal > 50%-50)/(signal > 50%-signal < 50%)xLN(conc > 50%/conc < 50%))).